Changes in the association of the catalytic subunit and the regulatory subunits of isozymes I and II of cAMP-dependent protein kinases (RI and RII, respectively) with the transcriptionally active chromatin fraction from rat liver were examined after a glucagon/theophylline injection and also after partial hepatectomy. Chromatin was partitioned into transcriptionally active and bulk, transcriptionally inactive fractions by digestion with micrococcal nuclease under appropriate conditions. In both experimental models, an increased content of catalytic and both RI and RII subunits was observed in chromatin fractions that were enriched in transcriptionally active DNA, particularly in the fraction associated with the residual nuclear matrix-lamina. The changes in the association of the subunits with these fractions paralleled the increases in intracellular cAMP levels and occurred in a time frame compatible with the changes in gene expression. The catalytic subunits could be removed from the nuclear matrix-lamina fraction by salt, whereas the two regulatory subunits remained tightly bound. The data support the concept of a direct role of the regulatory subunits of cAMP-dependent protein kinases in the induction of gene expression. However, we were unable to confirm that RII possessed an intrinsic topoisomerase activity.